Abstract

A homozygous mutation in a splice site of the CD81 gene was identified previously in a patient, as the cause in a case of common variable immune deficiency (CVID). CD19 expression is reduced in mice that lack CD81; however, B cells in this patient lacked completely CD19 surface expression. The mutation led to an absence of the CD81 protein on the cell surface and it was assumed that the CD81 protein was not produced. Here we demonstrate that a truncated human CD81 mutant (CD81mut) was actually produced, but retained intracellularly. We also demonstrate that the truncated CD81mut protein is in close proximity to the intracellularly sequestered CD19. However, this interaction does not enable normal CD19 maturation and surface expression. In addition, we show that specific domains of CD81 enable retrieval and trafficking of human CD19 to the cell surface. Finally, we demonstrate that surface expression of CD19 requires CD81, even in non-B cells.

Abstract

Hepatitis C Virus (HCV) infects 200 million individuals worldwide. Although several FDA approved drugs targeting the HCV serine protease and polymerase have shown promising results, there is a need for better drugs that are effective in treating a broader range of HCV genotypes and subtypes without being used in combination with interferon and/or ribavirin. Recently, two crystal structures of the core of the HCV E2 protein (E2c) have been determined, providing structural information that can now be used to target the E2 protein and develop drugs that disrupt the early stages of HCV infection by blocking E2's interaction with different host factors. Using the E2c structure as a template, we have created a structural model of the E2 protein core (residues 421-645) that contains the three amino acid segments that are not present in either structure. Computational docking of a diverse library of 1,715 small molecules to this model led to the identification of a set of 34 ligands predicted to bind near conserved amino acid residues involved in the HCV E2: CD81 interaction. Surface plasmon resonance detection was used to screen the ligand set for binding to recombinant E2 protein, and the best binders were subsequently tested to identify compounds that inhibit the infection of Huh-7 cells by HCV. One compound, 281816, blocked E2 binding to CD81 and inhibited HCV infection in a genotype-independent manner with IC50's ranging from 2.2 µM to 4.6 µM. 281816 blocked the early and late steps of cell-free HCV entry and also abrogated the cell-to-cell transmission of HCV. Collectively the results obtained with this new structural model of E2c suggest the development of small molecule inhibitors such as 281816 that target E2 and disrupt its interaction with CD81 may provide a new paradigm for HCV treatment.

Abstract

In B lymphocytes, the cell surface receptor CD38 is involved in apoptosis of immature B cells, proliferation and differentiation of mature B cells. Although CD38 has been establish as a receptor, its signaling has been only partially characterized. As a result of the lack of signaling motifs in the cytoplasmic domain, CD38 must use a co-receptor to induce signaling within the cell. Accordingly, CD38 has been associated with different receptors such as the T-cell receptor/CD3 complex on T cells, CD16 on natural killer cells and MHC class II molecules on monocytes. The CD19/CD81 complex has been proposed as a co-receptor for CD38 in human B lymphocytes, but little or no characterization has been performed in mice. In this study the contribution of the CD19/CD81 complex in murine CD38 signaling was evaluated. Proliferation assays were performed using CD19(-/-) or CD81(-/-) deficient mice; CFSE-labeled B lymphocytes from wild-type mice and CD19(-/-) , CD81(-/-) and CD38(-/-) deficient mice were stimulated with agonistic antibodies against CD38. Immunoprecipitation and immunofluorescence were also performed to detect protein-protein interactions. Our results indicate that the CD19/CD81 complex interacts with CD38 but this interaction is not required to induce proliferation in mouse B lymphocytes, suggesting that other receptors may contribute to the proliferation induced by CD38 in B lymphocytes.

Abstract

We have identified Tspan33 as a gene encoding a transmembrane protein exhibiting a restricted expression pattern including expression in activated B cells. TSPAN33 is a member of the tetraspanin family. TSPAN33 is not expressed in resting B cells, but is strongly induced in primary human B cells following activation. Human 2E2 cells, a Burkitt's lymphoma-derived B cell model of activation and differentiation, also upregulate TSPAN33 upon activation. TSPAN33 is expressed in several lymphomas including Hodgkin's and Diffuse large B cell lymphoma. TSPAN33 is also expressed in some autoimmune diseases where B cells participate in the pathology, including rheumatoid arthritis patients, systemic lupus erythematosus (SLE), and in spleen B cells from MRL/Fas(lpr/lpr) mice (a mouse model of SLE). We conclude that TSPAN33 may be used as a diagnostic biomarker or as a target for therapeutic antibodies for treatment of certain B cell lymphomas or autoimmune diseases.

Abstract

Bruton agammaglobulinemia tyrosine kinase (BTK) is a key protein in the B-cell receptor (BCR) signaling pathway and plays an essential role in the differentiation of B lymphocytes. X-linked agammaglobulinemia (XLA) is a primary humoral immunodeficiency caused by mutations in the gene encoding BTK. Previously, we identified two novel variations, L111P and E605G, in BTK; these are localized within the pleckstrin homology and Src homology 1 domains, respectively. In the present study, we evaluated the potential effects of these variations on the structural conformation and the function of BTK. Using in silico methods, we found that the L111P and E650G variations are not located directly in protein-protein interfaces but close to them. They distorted the native structural conformation of the BTK protein, affecting not only its geometry and stability but also its ability for protein recognition and in consequence its functionality. To confirm the results of the in silico assays, WT BTK, L111P, and E650G variants were expressed in the BTK-deficient DT40 cell line. The mutant proteins exhibited an absence of catalytic activity, aberrant redistribution after BCR-crosslinking, and deficient intracellular calcium mobilization. This work demonstrates that L111 and E605 residues are fundamental for the activation and function of BTK.

CD38 Through the Life of a Murine B LymphocyteIUBMB LIFEVences-Catalan, F., Santos-Argumedo, L.2011; 63 (10): 840-846

Abstract

CD38 is a 45 kDa transmembrane receptor expressed in B lymphocytes and other cells from the immune system. It is involved in apoptosis, cell activation, differentiation, and proliferation. CD38 has been used extensively to classify various subpopulations of lymphocytes in both humans and mice. It has also been used as a marker of poor prognosis in some lymphoid pathologies. However, CD38 is not a marker but rather an ectoenzyme and a receptor, where it performs several functions. The CD38 signaling pathway has only been partially studied in various cells of the immune system, where apparently the signaling is different depending on the lineage and differentiation state of the cell, leading to distinct outcomes. In this review, we provide an overview of well-established roles of CD38 signaling B lymphocytes from mice. We also discuss areas that need further clarification to get a broader image of how CD38 performs different functions in B cells and to understand its role in B lymphocyte biology under normal versus pathological conditions.